MicroRNAs (miRNA) are endogenously produced small non-coding regulatory RNA molecules. Through sequence complementation, miRNA interacts with specific messenger RNAs (mRNAs) and affects the stability of mRNAs and/or the progression of protein translation. It has been estimated that over 30% of mRNAs are regulated by miRNA. siRNA is a synthetic molecule, usually consisting of double-stranded RNA with short single-stranded ends, which is transfected into cells and binds to RISC. siRNA is perfectly complementary to a particular target mRNA and cleaves that target. siRNA also suppresses translation of many off-target mRNAs that have partial sequence complementarity. Because of their ability to regulate protein production, these small RNAs are being developed as therapeutics.
A significant number of microRNAs have been observed in the extracellular space. These extracellular miRNAs are stable and the changes in their spectrum have been demonstrated as sensitive and informative biomarkers for specific disease conditions. The existence of stable extracellular microRNAs also suggests the possibility of microRNA as one of the mediators involved in the cell-cell communication. In any event, the stability of microRNA in the extracellular space suggests a mechanism is available for stabilizing these inherently unstable molecules.
One possibility for such stabilization is complexation with proteins that are known to bind RNA. A number of such proteins are known; however, these have not been shown to protect RNA from degradation, and indeed, as shown below, not all RNA binding proteins are protective.
In addition to microRNAs as therapeutics, these molecules may be therapeutic targets as well. One possibility for attacking such targets is the use of small RNA complementary to miRNAs. For example, the mir122 miRNA, which is liver specific, is required for the replication of Hepatitis C virus. This is just one example of abnormal miRNA expression that is associated with disease where the pathogens can use pathogen-encoded miRNAs or utilize host miRNAs to modulate responses to the pathogens that benefit the pathogens themselves.
One of the major obstacles to use of small RNA including miRNA or siRNA as a therapeutic is the difficulty in delivery of stable RNA molecules into the body. In one approach, various chemical modifications of the nucleotide structures have been developed to prevent RNAse degradation and enhance stability. These modifications change the nature of the molecules and thus may affect the specific interaction with its intended target molecules and cause unpredictable adverse effects. Further, even though chemically modified RNA molecules are relatively stable in circulation, significant amounts are required to obtain the intended biological effects. Such high concentrations may generate immune responses preventing additional treatment based on similar molecules. In addition, the pharmacodynamic and pharmacokinetic properties of each of these modified RNA molecules need to be extensively investigated due to the unpredictable properties of adsorption, distribution, metabolism and excretion (ADME).
As noted above, a number of RNA binding proteins are known. It has been suggested that one of these NPM1 may be involved in shuttling RNAs and ribosomal proteins to the cytosol (Leask, A., J. Cell Commun. Signal (2009) 3:85-86) and in a recent report it has also been identified outside the cell (Nawa, Y., et al., J. Leukoc. Biol. (2009) 86:1-9).